A major limitation in the toxicity of chimeric toxins is the efficiency with which enzymically active portions of toxin moieties are transferred into the cytosolic compartment of susceptible cells. Our primary goals are to elucidate features of diphtheria and Pseudomonas toxins that function in the transfer and to build a technological base for using molecular genetics to alter these toxin moieties in desired manners. We have recently cloned into E. coli a diphtheria toxin gene fragment that encodes the toxin leader peptide, the A fragment, and the hydrophobic portion of B and have also obtained a cloned form of the Pseudomonas toxin gene. These are now being subjected to site-directed mutagenesis in order to probe structure/function relationships in the two toxins, including catalytic activities and membrane transport functions. Concurrent studies on the protein level, including lipid bilayer studies and binding and transport studies in tissue culture, are being undertaken in order to provide a basis for understanding in greater detail the limiting factors in the transfer of enzymically active portions of toxin moieties into the cytosolic compartment. (HI)